Transcription induced byrole dsRNA data of natural analysis formation. antisense of overlapping Instead, transcripts itgene suggests pairs in Arabidopsis alternative in Arabidopsis roles for thaliana dsRNAs argues such as regulation againstofa alternative predominant splicing RNA degradation in polyade-
Conclusion: The results argue against a predominant RNA degradation effect induced by dsRNA formation. Instead, our data support alternative roles for dsRNAs. They suggest that at least for a subgroup of COPs, antisense expression may induce alternative splicing or polyadenylation.
teria, the frequencies for overlapping gene pairs vary between 4% and 9% for the human genome, 1.7%-14% in the murine genome, and up to 22% in the fly genome [1]. The predominant composition of overlapping gene pairs is an antiparallel
Genome Biology 2005, 6:R51
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Genome-wide searches in the genomes of several species have identified a surprisingly high proportion of overlapping gene pairs. Depending on the sample sizes analyzed and search cri-
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Results: We have examined transcription data for overlapping gene pairs in Arabidopsis thaliana. On the basis of an analysis of transcripts with coding regions, we find the majority of overlapping gene pairs to be convergently overlapping pairs (COPs), with the potential for dsRNA formation. In all tissues, COP transcripts are present at a higher frequency compared to the overall gene pool. The probability that both the sense and antisense copy of a COP are co-transcribed matches the theoretical value for coexpression under the assumption that the expression of one partner does not affect the expression of the other. Among COPs, we observe an over-representation of spliced (intron-containing) genes (90%) and of genes with alternatively spliced transcripts. For loci where antisense transcripts overlap with sense transcript introns, we also find a significant bias in favor of alternative splicing and variation of polyadenylation.
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Background: Overlapping transcripts in antisense orientation have the potential to form doublestranded RNA (dsRNA), a substrate for a number of different RNA-modification pathways. One prominent route for dsRNA is its breakdown by Dicer enzyme complexes into small RNAs, a pathway that is widely exploited by RNA interference technology to inactivate defined genes in transgenic lines. The significance of this pathway for endogenous gene regulation remains unclear.
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The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2005/6/6/R51
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Addresses: *School of Biochemistry and Microbiology, University of Leeds, Leeds LS2 9JT, UK. †Centre for Plant Science, The University of Leeds, Leeds LS2 9JT, UK.
R51.2 Genome Biology 2005,
Volume 6, Issue 6, Article R51
Jen et al.
convergent arrangement [2,3], where sense and antisense genes overlap within their 3' regions. Joint expression of both these genes in the same cell would allow the partly overlapping transcripts to associate as dsRNA molecules, which may interfere with RNA processing, transport, stability or other molecular mechanisms. Convergently overlapping gene pairs (COPs) can therefore provide the source for natural antisense transcripts (NATs) that may act as regulators of the sense gene. In addition to NATs being transcribed from the same locus as the sense transcript (cis-NATs), NATs can be transcribed from a different locus (trans-NATs), as illustrated by a search for overlapping transcripts with coding capacity in the human genome, which identified 87 cis-NATs and 80 trans-NATs [3]. In bacteria, more than 100 NATs are involved in the regulation of a variety of biological functions, including the control of copy number, conjugation and post-segregational killing in plasmids, lysis/lysogeny switches in phages, and transposition frequency in transposons [4]. In eukaryotes, a very detailed characterization of the molecular role of specific NATs has only been achieved for a few examples. NAT-mediated interference with splicing is illustrated by the alternative processing of mRNAs of the gene for the thyroid hormone receptor ErbAα, which is regulated by an antisense transcript [5]. Overlapping genes can share a bidirectional poly(A) region as demonstrated for the human genes ABHD1 and Sec12 [6]. Several examples document the fact that antisense transcripts can increase sense transcript stability, when dsRNA regions cover the 3' untranslated region (UTR) and possibly mask out target sequences for RNA cleavage [7]. Alternatively, RNA duplex formation can increase transcript sensitivity and induce site-specific cleavage, as shown for the human TYMS mRNA and TRS antisense transcripts [8]. An example of RNA interference (RNAi)-based regulation of an endogenous gene via NATs is the repression of the testisexpressed Stellate gene in Drosophila by paralogous Su(Ste) tandem repeats [9]. Both strands of repressor Su(Ste) repeats are transcribed, producing sense and antisense RNA, most probably as part of a dsRNA-based silencing mechanism, as Stellate silencing is associated with the presence of short Su(Ste) RNAs. Antisense expression can also affect translation, as illustrated by the influence of an antisense transcript on the translation of different isoforms of fibroblast growth factor-2 (FGF2) [10]. In the nucleus, dsRNA can be edited by dsRNA-dependent adenosine deaminases, which convert about 50% of adenosine residues into inosines, leading to the unwinding of the RNA duplex [11]. Inosine-containing RNAs are not translated as they are retained in the nucleus [12].
http://genomebiology.com/2005/6/6/R51
transcript annotation allows such predictions. There is some evidence for an over-representation of overlapping genes among specific functional categories, that is, imprinted genes and DNA repair genes [1]. Twenty-two out of 58 known imprinted murine genes are transcribed from both strands. Frequently, one partner transcribes a noncoding RNA. Antisense transcripts may regulate imprinting states of the sense promoter (Kcnq1/Kcnq1ot1) or may induce dsRNA-based gene silencing as proposed for Ifd2R/Air. About 20% of known human DNA repair genes overlap either convergently or divergently in an antiparallel arrangement [1]. Mammalian mRNAs that form sense-antisense pairs frequently exhibit reciprocal expression patterns, but permanent coexpression of sense and antisense transcripts can also occur in some tissues, although it is difficult to prove that both genes are transcribed in the same cell. Coexistence of sense and antisense transcripts may indicate a stabilizing effect of dsRNA, or it may depict cases where RNA duplex formation is impaired as a result of secondary structures, or because sense and antisense transcripts or the enzymes required for duplex formation are separated by compartmentalization [13]. To gain an insight into the existence and role of overlapping antisense pairs in plants, we have screened the Arabidopsis thaliana genome for COPs with sense and antisense genes that encode a protein, and have compared the expression profile of the associated genes. All types
Sense-antisense orientation A
